AIM:Survivin, a recently identified member of the inhibitor of apoptosis protein family, is expressed during development and in various human cancers. However, its expression in normal tissues and clinical relevance i...AIM:Survivin, a recently identified member of the inhibitor of apoptosis protein family, is expressed during development and in various human cancers. However, its expression in normal tissues and clinical relevance in cancers are still debated. In the present study, we analyzed the expression of the survivin gene in human primary and metastatic gastric cancer cells as well as in paired epithelial cells from normal gastric mucosa by means of a novel laser capture microdissection (LCM) technique coupled with reverse transcription -polymerase chain reaction (RT-PCR).METHODS: Thirty patients who had undergone gastrectomy with lymph node dissection for gastric cancer without preoperative treatments were included. Neoplastic tissue, metastatic lymph nodes, and apparently uninvolved normal tissue were collected from each patient. LCM-captured 'pure' cell groups were respectively subjected to RT-PCR analysis with primers specific for the survivin gene.RESULTS: Of the paired samples from 30 gastric cancer patients studied, 24 (80%) primary gastric cancer cell groups and 7 (23%) adjacent morphologically 'normal' gastric epithelial cell groups were shown to have a detectable survivin expression. There was a statistically significant difference in suvivin expression between these two groups (P<0.01). Meanwhile, 95% (19/20) of the metastatic gastric cancer cell groups from lymph nodes had a clear expression of the survivin gene. However, no significant correlation between survivin expression and clinicopathological features of gastric cancer was observed in the present study. CONCLUSION: Survivin expression is present in the majority of gastric cancer cell groups obtained by LCM techniques. The high expression rate in metastatic lesions suggests a possible role of survivin in cancer invasiveness and metastasis. It may contribute to the detection of gastric cancer micrometastasis as a potential molecular marker. In addition, the high expression percentage renders survivin a potential target in the therapy for gastric cancer.展开更多
AIM: To verify the effectiveness of denaturing highperformance liquid chromatography (DHPLC) in detecting somatic mutation of p53 gene in gastric carcinoma tissues.The superiority of this method has been proved in the...AIM: To verify the effectiveness of denaturing highperformance liquid chromatography (DHPLC) in detecting somatic mutation of p53 gene in gastric carcinoma tissues.The superiority of this method has been proved in the detection of germline mutations, but it was not very affirmative with respect to somatic mutations in tumor specimens. ST7 gene, a candidate tumor suppressor gene identified recently at human chromosome 7q31.1, was also detected because LOH at this site has also been widely reported in stomach cancer.METHODS: DNA was extracted from 39 cases of surgical gastric carcinoma specimen and their correspondent normal mucosa. Seven fragments spanning the 11 exons were used to detect the mutation of p53 gene and the four exons reported to have mutations in ST7 gene were amplified by PCR and directly analyzed by DHPLC without mixing with wild-type allele.RESULTS: In the analysis of p53 gene mutation, 9 aberrant DHPLC chromatographies were found in tumor tissues, while their normal-adjacent counterparts running in parallel showed a normal shape. Subsequent sequencing revealed nine sequence variations, 1 polymorphism and 8 mutations including 3 mutations not reported before. The mutation rate of p53 gene (21%) was consistent with that previously reported. Furthermore, no additional aberrant chromatography was found when wild-type DNA was added into the DNA of other 30 tumor samples that showed normal shapes previously. The positivity of p53 mutations was significantly higher in intestinal-type carcinomas (40 %) than that in diffuse-type (8.33 %) carcinomas of the stomach. No mutation of ST7 gene was found.CONCLUSION: DHPLC is a very convenient method for the detection of somatic mutations in gastric carcinoma. The amount of wild type alleles supplied by the non-tumorous cells in gastric tumor specimens is enough to form heteroduplex with mutant alleles for DHPLC detection. ST7 gene may not be the target gene of inactivation at 7q31 site in gastric carcinoma.展开更多
Objective To evaluate the prevalence of rhodopsin (RHO) mutations and the genotype-phenotype relationships in Chinese patients with autosomal dominant retinitis pigmentosa (ADRP) by conformation sensitive gel electrop...Objective To evaluate the prevalence of rhodopsin (RHO) mutations and the genotype-phenotype relationships in Chinese patients with autosomal dominant retinitis pigmentosa (ADRP) by conformation sensitive gel electrophoresis (CSGE) and direct DNA sequencing. Methods We have screened the five coding exons and splice sites of RHO gene in 27 probands who had no relativity from Chinese ADRP families and 100 normal controls to identify disease-associated mutations, using CSGE and direct DNA sequencing. Family members of some probands with disease-associated mutations were also genotyped to determine whether the RHO mutations segregated with retinitis pigmentosa (RP) in their families. Results Two RHO mutations, Pro347Leu and Pro327 (1-bp del), were identified separately in two families, thus the frequency of RHO mutations among this set of Chinese ADRP families is about 7.4% (2/27). Pro347Leu mutation was found in one ADRP proband as well as three her children who also had RP. She had relatively early onset at about 17 years. The only one child without this mutation had no symptom or sign of RP at age of 34. Pro327 (1-bp del) was identified in a late-onset ADRP patient, who appeared night blindness around 30 years old and in her fifties electroretinogram (ERG) has been flat in both scotopic and photopic phases. Family analysis showed that this mutation also existed in her younger dau-ghter and her elder sister, both of them also had RP. Three other family members were genotypically and phenotypically normal. Neither of the two mutations was detected in 100 normal controls.Conclusions The frequency of RHO mutations in Chinese patients was lower than that in Europe and North America. The phenotype of the patients with Pro347Leu corresponded to type 1 ADRP, with severe rod degeneration and some cone preservation later, while the phenotype of the patients carrying Pro327 (1-bp del) corresponded to type 2 ADRP, with a concomitant loss of rod and cone visual function. CSGE was found to be a sensitive, simple, and practical method for the screening of a large number of samples under highly reproducible conditions, and could be utilized in routine molecular diagnostic laboratories.展开更多
AIM: To study the effect of cyclin G2 on proliferation of gastric adenocarcinoma cell line-SGC-7901 cell in vitro.METHODS: By use of cation lipofectamine transfection reagent,the pIRES-G2 and pIRESneo plasmids were tr...AIM: To study the effect of cyclin G2 on proliferation of gastric adenocarcinoma cell line-SGC-7901 cell in vitro.METHODS: By use of cation lipofectamine transfection reagent,the pIRES-G2 and pIRESneo plasmids were transferred into SGC-7901cell line. Anticlones were selected by G418. Positive clones were observed and counted using Giemsa staining.Cell proliferative ability was assayed by Ml-r.RESULTS: (1) The done number of pIRES-G2 group decreased,clone volume reduced. The number of cell clones in pIRESneo group was 87+3, that of pIRES-G2 group was 53+4,occupying 60.1% of pIRESneo group, there was significant difference obviously (P<0.01, t=15.45). (2) The average absorbance of clone cell obtained by stable transfection of pIRES-G2 at 570 nm was 1.6966+0.2125, the average absorbance of clone cell obtained by stable transfection of pIRESneo at 570 nm was 2.1182+0.3675, there was significant difference between them (P<0.01, t=3.412).CONCLUSION: Cyclin G2 can inhibit SGC-7901cell proliferative ability obviously, it may be a negative regulator in cell cycle regulation.展开更多
Objective Intra-operative coagulopathy has a close relationship with blood loss and the prognosis of patients. Rapid-thrombelastography （r-TEG） is a comprehensive assessment of coagulation abnormalities and also an...Objective Intra-operative coagulopathy has a close relationship with blood loss and the prognosis of patients. Rapid-thrombelastography （r-TEG） is a comprehensive assessment of coagulation abnormalities and also an effective way for constructing blood transfusion. This study attempts to investigate the correlation of r-TEG indices with intra-operative hemorrhage. Methods Patients who underwent transphenoidal hypophysectomy and craniotomy from January 15 to April 30, 2013 in Peking Union Medical College hospital were recruited. All patients had pre- and post-operative r-TEG and conventional coagulation tests （CCTs）. Patients’ information and intra-operative blood loss as a percentage of estimated blood volume were recorded. Spearman’s correlation analyses were used for discovering the relationship between indices in r-TEG or CCTs and the intra-operative blood loss. The significant correlated index of r-TEG was further investigated using linear regression analysis. Results A total of 181 patients participated in this study. Intra-operative change of α-angle, which reflects the fibrinogen level and function, was the only r-TEG index that correlated with blood loss significantly （P=0.013, r= ？0.184）, thus challenging the current empirical cognition of the effects of intra-operative hemorrhage on coagulation. As intra-operative blood loss increased, α-angle decreased, and every 1% loss of estimated blood volume （EBV） led to 0.60 degree decrease of α-angle. As for CCT results, changes of fibrinogen and platelet count were also significantly correlated with blood loss （P=0.015 and P=0.001, respectively）.Conclusions Peri-operative change of α-angle, as an index of r-TEG, exhibited a significant negative correlation with intra-operative blood loss. The impact of hemorrhage on fibrinogen, instead of clotting factors, should be scrutinized.展开更多
基金Supported by the Nationa 1973 Program of China,No.G 1998051203,the National Science Fund for Distinguished Young Scholars of China,No.30125017,and the MOE TRAPOYT Program of China,No.1999-96
文摘AIM:Survivin, a recently identified member of the inhibitor of apoptosis protein family, is expressed during development and in various human cancers. However, its expression in normal tissues and clinical relevance in cancers are still debated. In the present study, we analyzed the expression of the survivin gene in human primary and metastatic gastric cancer cells as well as in paired epithelial cells from normal gastric mucosa by means of a novel laser capture microdissection (LCM) technique coupled with reverse transcription -polymerase chain reaction (RT-PCR).METHODS: Thirty patients who had undergone gastrectomy with lymph node dissection for gastric cancer without preoperative treatments were included. Neoplastic tissue, metastatic lymph nodes, and apparently uninvolved normal tissue were collected from each patient. LCM-captured 'pure' cell groups were respectively subjected to RT-PCR analysis with primers specific for the survivin gene.RESULTS: Of the paired samples from 30 gastric cancer patients studied, 24 (80%) primary gastric cancer cell groups and 7 (23%) adjacent morphologically 'normal' gastric epithelial cell groups were shown to have a detectable survivin expression. There was a statistically significant difference in suvivin expression between these two groups (P<0.01). Meanwhile, 95% (19/20) of the metastatic gastric cancer cell groups from lymph nodes had a clear expression of the survivin gene. However, no significant correlation between survivin expression and clinicopathological features of gastric cancer was observed in the present study. CONCLUSION: Survivin expression is present in the majority of gastric cancer cell groups obtained by LCM techniques. The high expression rate in metastatic lesions suggests a possible role of survivin in cancer invasiveness and metastasis. It may contribute to the detection of gastric cancer micrometastasis as a potential molecular marker. In addition, the high expression percentage renders survivin a potential target in the therapy for gastric cancer.
基金National Science Fund for Distinguished Young Scholars,No.30125017the Major State Basic Research Development Program of China(973 Program),No.G1998051203
文摘AIM: To verify the effectiveness of denaturing highperformance liquid chromatography (DHPLC) in detecting somatic mutation of p53 gene in gastric carcinoma tissues.The superiority of this method has been proved in the detection of germline mutations, but it was not very affirmative with respect to somatic mutations in tumor specimens. ST7 gene, a candidate tumor suppressor gene identified recently at human chromosome 7q31.1, was also detected because LOH at this site has also been widely reported in stomach cancer.METHODS: DNA was extracted from 39 cases of surgical gastric carcinoma specimen and their correspondent normal mucosa. Seven fragments spanning the 11 exons were used to detect the mutation of p53 gene and the four exons reported to have mutations in ST7 gene were amplified by PCR and directly analyzed by DHPLC without mixing with wild-type allele.RESULTS: In the analysis of p53 gene mutation, 9 aberrant DHPLC chromatographies were found in tumor tissues, while their normal-adjacent counterparts running in parallel showed a normal shape. Subsequent sequencing revealed nine sequence variations, 1 polymorphism and 8 mutations including 3 mutations not reported before. The mutation rate of p53 gene (21%) was consistent with that previously reported. Furthermore, no additional aberrant chromatography was found when wild-type DNA was added into the DNA of other 30 tumor samples that showed normal shapes previously. The positivity of p53 mutations was significantly higher in intestinal-type carcinomas (40 %) than that in diffuse-type (8.33 %) carcinomas of the stomach. No mutation of ST7 gene was found.CONCLUSION: DHPLC is a very convenient method for the detection of somatic mutations in gastric carcinoma. The amount of wild type alleles supplied by the non-tumorous cells in gastric tumor specimens is enough to form heteroduplex with mutant alleles for DHPLC detection. ST7 gene may not be the target gene of inactivation at 7q31 site in gastric carcinoma.
文摘Objective To evaluate the prevalence of rhodopsin (RHO) mutations and the genotype-phenotype relationships in Chinese patients with autosomal dominant retinitis pigmentosa (ADRP) by conformation sensitive gel electrophoresis (CSGE) and direct DNA sequencing. Methods We have screened the five coding exons and splice sites of RHO gene in 27 probands who had no relativity from Chinese ADRP families and 100 normal controls to identify disease-associated mutations, using CSGE and direct DNA sequencing. Family members of some probands with disease-associated mutations were also genotyped to determine whether the RHO mutations segregated with retinitis pigmentosa (RP) in their families. Results Two RHO mutations, Pro347Leu and Pro327 (1-bp del), were identified separately in two families, thus the frequency of RHO mutations among this set of Chinese ADRP families is about 7.4% (2/27). Pro347Leu mutation was found in one ADRP proband as well as three her children who also had RP. She had relatively early onset at about 17 years. The only one child without this mutation had no symptom or sign of RP at age of 34. Pro327 (1-bp del) was identified in a late-onset ADRP patient, who appeared night blindness around 30 years old and in her fifties electroretinogram (ERG) has been flat in both scotopic and photopic phases. Family analysis showed that this mutation also existed in her younger dau-ghter and her elder sister, both of them also had RP. Three other family members were genotypically and phenotypically normal. Neither of the two mutations was detected in 100 normal controls.Conclusions The frequency of RHO mutations in Chinese patients was lower than that in Europe and North America. The phenotype of the patients with Pro347Leu corresponded to type 1 ADRP, with severe rod degeneration and some cone preservation later, while the phenotype of the patients carrying Pro327 (1-bp del) corresponded to type 2 ADRP, with a concomitant loss of rod and cone visual function. CSGE was found to be a sensitive, simple, and practical method for the screening of a large number of samples under highly reproducible conditions, and could be utilized in routine molecular diagnostic laboratories.
基金Supported by a grant for Distinguished Young Teachers of HigherEducation of the Ministry of Education of the Teaching and ResearchEncouragement Plan
文摘AIM: To study the effect of cyclin G2 on proliferation of gastric adenocarcinoma cell line-SGC-7901 cell in vitro.METHODS: By use of cation lipofectamine transfection reagent,the pIRES-G2 and pIRESneo plasmids were transferred into SGC-7901cell line. Anticlones were selected by G418. Positive clones were observed and counted using Giemsa staining.Cell proliferative ability was assayed by Ml-r.RESULTS: (1) The done number of pIRES-G2 group decreased,clone volume reduced. The number of cell clones in pIRESneo group was 87+3, that of pIRES-G2 group was 53+4,occupying 60.1% of pIRESneo group, there was significant difference obviously (P<0.01, t=15.45). (2) The average absorbance of clone cell obtained by stable transfection of pIRES-G2 at 570 nm was 1.6966+0.2125, the average absorbance of clone cell obtained by stable transfection of pIRESneo at 570 nm was 2.1182+0.3675, there was significant difference between them (P<0.01, t=3.412).CONCLUSION: Cyclin G2 can inhibit SGC-7901cell proliferative ability obviously, it may be a negative regulator in cell cycle regulation.
文摘Objective Intra-operative coagulopathy has a close relationship with blood loss and the prognosis of patients. Rapid-thrombelastography （r-TEG） is a comprehensive assessment of coagulation abnormalities and also an effective way for constructing blood transfusion. This study attempts to investigate the correlation of r-TEG indices with intra-operative hemorrhage. Methods Patients who underwent transphenoidal hypophysectomy and craniotomy from January 15 to April 30, 2013 in Peking Union Medical College hospital were recruited. All patients had pre- and post-operative r-TEG and conventional coagulation tests （CCTs）. Patients’ information and intra-operative blood loss as a percentage of estimated blood volume were recorded. Spearman’s correlation analyses were used for discovering the relationship between indices in r-TEG or CCTs and the intra-operative blood loss. The significant correlated index of r-TEG was further investigated using linear regression analysis. Results A total of 181 patients participated in this study. Intra-operative change of α-angle, which reflects the fibrinogen level and function, was the only r-TEG index that correlated with blood loss significantly （P=0.013, r= ？0.184）, thus challenging the current empirical cognition of the effects of intra-operative hemorrhage on coagulation. As intra-operative blood loss increased, α-angle decreased, and every 1% loss of estimated blood volume （EBV） led to 0.60 degree decrease of α-angle. As for CCT results, changes of fibrinogen and platelet count were also significantly correlated with blood loss （P=0.015 and P=0.001, respectively）.Conclusions Peri-operative change of α-angle, as an index of r-TEG, exhibited a significant negative correlation with intra-operative blood loss. The impact of hemorrhage on fibrinogen, instead of clotting factors, should be scrutinized.
